Rifamycin b hydrazides



3,197,468 United States Patent Office m, M, ,7, 1,65

istry and Biochemistry of Fungi and Yeasts Congress RIF clgi ggflfgnRAzmEs which was held in Dublin on July 18, 1963), which estab- AMYl' h d f v Piero Sensi and Nicola Maggi, Milan, Italy, assignors to e ornfamycm B the followm Structure Lepetit S.p.A., Milan, Ital y M M H NoDrawing. Filed Feb. 5, 1964, Ser. No. 342,804 e e Claims priority,application Great Britain, Feb. 18, 1963, 6,460/ 63 16 Claims. (Cl.260239.3)

The present invention concerns new antibiotic substances and the processfor their preparation. In the US. copencling application serial No.12,654, filed on March 4, 1960, now U.S. Patent No. 3,150,046, thepreparation M60 of the antibiotic rifamycin by fermentation of a strainof Str. mediterranei ATCC 13685 is described. As stated j in saidapplication, rifamycin is a mixture of antibiotic substances. One ofthese substances, rifamycin B, having 0 the crude formula O l-1 N0 is adiacid (pH 1 /2=2.8, pH 2%:61) and one of the acidic groups is acarboxyl group. One of the particular properties of this antibiotic Theabove formula explains of course how rifamycins a m jlyD" O sto a in e ais that it increases in activity when dissolved in water, i.e. O, S andSV are produced from rifamycin B. The seit is converted into anothersubstance having higher antiquence of such modifications may beillustrated by conbacterial activity. The more active product, calledrisidering the sole naphthalene ring present in the molefamycin S, hasthe crude formula C H NO and by cule:

OH EH1 O H E) O H O l] l T M NH Oxid. Me NH Hydrolysis M s i l- H l- H Hl 1 ll OCHnCOOH ""0 0 H CH:& 0

rifamycin B rifamycin O riiamycin S l1/ Reducing agents rifarnycin SVmild reduction can be converted into another new anti- The presentinvention concerns new derivatives of ribiotic of the rifamycin class,rifamycin SV (C31H47NO12)- famycin B in which the carboxyl group isblocked by The Process for Prepaflng rlfamycm 3V ComprisesOXldaconversion into hydrazides. The process for preparing the tion ofIifamycin B m rifamycin hydrolysis of Iifamy new derivatives consists incondensing rifamycin B with i'g g gg g gi gf i g gigi ii gg fisubstituted hydrazines in the presence of dehydrating S and fifamycin Svlack the carboxyl group which is Set agents such as carbodnnndes in aninert anhydrous solfree in the form of glycolic acid during theactivation step. Vent- Th6 Pmferred Condensing agent 15 dleyclohexylcarThe structure of rifarnycins has been recently elucidated bodiimide, thePmferred Solvent is tetrahydmfuran- The by Prelog and co-workers(communication at the Cherncondensation is preferably carried out atroom tempera- 71 5 5 55 8 55 50 758944977 999997800 %ms4fiwmwllmflumlo2mmw 1mo1.1000033500000000 in healing rium tuberculosis as t somemicroorganisms and the high activity zides.

ing the toxicity in rats flective doses f the described rifamycin BToxicity in the rat, LDsmi.

mgJkg.

7 8 n G 48577 697553555 21 4 O 7 Umyt S 72%3224521662 62 63 33W5 h k 1 Nk r D. .g na Dm am 1 rm E WN w 53 6 56 05 n O 7 672045940 5420 0 .mwm%%%142222122N7218 I O o a 0 D D m E v. 000802082202299550010059 01 747120465638984 tal infection by staphylococci have ty of rifamycin Bhydra ions agams sis are given.

The following data show drazides possess a hi h antibiotic act1v1tyagainst grampositive bacteria and Mycobacte shown in the followingtable, where the minimal inhibitory concentrat M. tuberculoPharmacological tests concern rats with experimen been carried out onseveral 0 hydrazides. and the low toxic Riiamycin B hydrazides withTABLE Minimal inhibitory concentrations /ml.

0. M 00 00 00 QQQQQQQQLQQQQQQQQ00Q00100000000 000000000 000000000000555500055555000 H 000000555= v000005555555222200022222565 0 22222011222 11. d U 6 H 5 5 555 127 w 00 0 5 000 5 5555555000 .5 .52555 .Dmmmo0m0 2mnm 2 000 222222255002021222 Ru 222222 212 1 m my 2155522222222222222 52225222 .1 0000006 L 2 6 0006 0 6 566600666525602666000000 3 1 000 6 1 1 6 222222 212 2 2255 5 55252 5 5 00 00 005552262605025255555 W 00mm00 HHm00mHm22211 1 52012122277 222222 222 1 mH 55 5 55 5 55 92 2299 457 46443555 M 1525 mmMMwmll500mM80000073500000777 W omLHwEJOHQQQQQQRWKWMQQQQQQQQQQQUU100000000 555 5 2 5 2 5124752 5 123171 715 75 534.8 0 0000 1 00 0001111000212122089000125OWOOOOWOBOOMOOOOOOO000000000000 sparingly soluble The concentratedS. faccalis S. haemol.

. 5 32 1 23 821n O u118h0 2 5 1L 1 5 0200 0M07550000000000010 00000011117522L0QQQQQQLMQQQQQQQQQQQQQOU0000000 In particular the M. auraus fpetroleum ether the crude 40 and the per es and subcutaneous e in Bhydrazide having the tetrahydrofuran, crystallises out. solution isdiluted with dilute hydrochloric acid or sulphuric acid and extractedseveral times with ethyl acetate.

Rifamyein B hydrazlde withture (20-25 C.) in 1-3 hours. Afterconcentration of the solvent the formed dicyclohexylurea, 1n

hydrazide precipitates. The hydrazides are recrystallised frombenzene-dicyclohexane or carbon tetrachloride.

Accordingly, the present invention provides as a novel compound ahydrazide of rifamycin B.

tion provides a rifamyc general formula:

m n n 9 u m .mammnm mmemmmmmmmmm i .i wr mmnmmmmmmmmmoh nmmm m m mmm mymwmmmmm mmnwm h v. dh v. r lvsh h 1v. h mo pp h ml h hlyh o m ipi l y hl ly h mo pO otypyhlvltypyhlyt. on iono t eho y in ln tr o tru omm mi mmi mamwmfimmammmeammmwasm nwn ynnmm ynnnnnnm nw nw y hhhhyy yppppyyy e omeh m aaaemmmnwmmmmmmnmem mmm MMMMEEEDPPPPBBB n44441111 K0 0 F6 Ru 6 G 3-6 w h mwawwn h w 1 m mm m vfi w n T eh if 23 a h 6 HS 5 2d R11, MN. MWtd r m u S h e e r% e t k I Hi m a f t. O S 0 d fdnna ss M a fi onhe netb a .a w. m em c m r, m h n e m Hm c a w. e r aOnmemYi b W uuux m m s,Wh m o g e o m g d ED. 0 b u e mumsm W.m w. r 0mr 0 d oh fl r a a v. etew rP wm wm h f y 0 n D. m wn ed m a m e re n c h a m m m a a a n m C.l rfon um wwm mmao m w o 4 mm a g s o a n r 2 a t d y Rmmmwbum ow m mgfi dmWSa C a d H g S 2 .1 n f D m H S m y a u 0..l n p m mm mwm w 1 y tb C n f a h ce 0.1 hO fm mmnmwm iwm m wfaa emi am colvc Y m s nror m lnbt n u r c P lfi hwb hm mmn smr fl d mih afl .1e m w n mw n w wcmama hddmmpnsw The ri h ethyl acetate is then concentrated to a small volume andby the addition 0 rifamycin inven mples are illustrative EXAMPLE 1Rifamycin B trimethylhydrazide An amount of 5 g in B is suspended l Then1.350 g. of dicyclohexyl- The following non-limitative exa invention.

of rifamyc of tetrahydrofuran of the in lues at 220-225 m 304-306 mg and420-425 mp. in buffer solut specific extinctions ion at pH 7.3. TheirThe rifamycin B hy- 7 m 5 carbodiimide are added followed by 0.500 g. oftrimethylhydrazine dissolved in 25 ml. of tetrahydrofuran. After theaddition, carried out in 30 minutes under strong stirring, the mixtureis allowed to stand for two hours, then concentrated to 3040 ml. Theprecipitated dicyclohexylurea is filtered off, the filtrate is pouredinto dilute hydrochloric acid and extracted with ethyl acetate. Theethyl acetate extract is dried over sodium sulphate, filtered,concentrated to 30-40 ml. and poured into 500 ml. of petroleum ether.The hydrazide precipitates in the form of a yellow orange amorphouspowder which is collected and dried in vacuo. Yield 4.5 g. The crudeproduct is dissolved in 2500 ml. of cyclohexane at 60 C., filtered fromsome insoluble portion and concentrated to about 100 ml. Thesemi-crystalline hydrazide precipitated during the concentration iscollected, giving 2.6 g. of rifamycin B trimethylhydrazide.Analysis.Calcd. for C H N O :C, 65.67; H, 6.41; N, 4.68; M.W. 895.970.Found: C, 62.27; H, 7.31; N, 4.80.

EXAMPLE 2 Rifamycin B trimethylhydrazide sodium salt Five grams ofrifamycin B triethylhydrazide are dissolved in .100 ml. of methanol andthe solution is adjusted to pH 8.6 with sodium methoxide. An amount of100 ml. of methyl isobutyl ketone is added, the solution is concentratedto a small volume in vacuo and the concentrate is poured into 500 ml. ofethyl ether. The precipitated rifamycin B trimethylhydrazide sodium saltis then collected and dried in vacuo. It appears as a yellowish powder,very soluble in water, fairly soluble in methanol and acetone andsparingly soluble in ethyl ether and petroleum ether.

EXAMPLES 3-13 According to the process of Example 1 the followingrit'amycin B hydrazides were prepared. Their corresponding sodium saltswere also prepared according to the process of Example 2.

Condensation product of rifamycin B with phenylhydra- Zine: C H N O M.W.845.914. Analysis: C, 63.88; H, 6.5; N, 4.96. Found: C, 63.13; H, 6.94;N, 5.10.

Condensation product of rifamycin B with p-phenylmethylhydrazine: C H NO M.W. 859.940. Analysis: C, 64.19; H, 6.67; N, 4.88. Found: C, 63.3; H,6.98; N, 4.94.

Condensation product of rifamycin B with (4-pyridyl)- carbohydrazine: CH N O M.W. 874.914. Analysis: C, 61.71; H, 6.21; N, 6.39. Found: C,60.40; H, 6.59; N, 6.40.

Condensation product of rifamycin B with (4-pyridyl)- carbohydrazidine:C H N O M.W. 873.930. Analysis: C, 61.82; H, 6.34; N, 8.01. Found: C,60.79; H, 6.48; N, 8.32.

Condensation product of rifamycin B with 4-aminomorpholine: C43H57N3O14,Analysis: C, 61.43; H, 6.83; N, 5.00. Found: C, 60.04; H, 6.24; N, 4.77.

Condensation product of rifamycin B with aminothiou- Tea: C40H52N4SO13,Analysis: C, H, 6.32; N, 6.75; S, 3.86. Found: C, 57.56; H, 6.79; N,6.52; S, 3.17.

Condensation product of rifamycin B with dimethylethylhydrazine: C H N OM.W. 825.95. Analysis: C, 62.48; H, 7.19; N, 5.08. Found: 61.62; H,6.34; N, 4.65.

Condensation product of rifamycin B with dimethylpropylhydrazine: C H NO M.W. 839.972. Analysis: C, 62.88; H, 7.31; N, 4.99. Found: C, 62.63;H, 7.67; N, 4.92.

Condensation product of rifamycin B with dimethylbutylhydrazine: C H N OM.W. 853.978. Analysis: C, 63.23; H, 7.42; N, 4.91. Found: C, 63.19; H,7.52; N, 4.71.

Condensation product of rifamycin B with dimethylamylhydrazine: C H N OM.W. 868.004. Analysis: C, 63.64; H, 7.54; N, 4.84. Found: C, 63.53; H,7.61; N, 4.99.

Condensation product of rifamycin B with diethylmethylhydrazine: C H N OM.W. 839.952. Analysis: C, 62.88; H, 7.31; N, 5.00. Found: C, 63.00; H,7.60; N, 4.89.

EXAMPLES 14-35 Condensation product of rifamycin B with hydrazine:C39'H51N3013, M.W. 769.8. Analysis: C, 60.84; H, 6.67; N, 5.45. Found:C, 61.12; H, 7.20; N, 5.80.

Condensation product of rifamycin B with di, n-propylethyl hydrazine: CH N O M.W. 813.6. Analysis: C, 60.6; H, 6.77; N, 5.16. Found: C, 61.02;H, 7.15; N, 5.

Condensation product of rifamycin B with 1-amino-2,4- dioxo imidazoline:C H N O M.W. 852.3. Analysis: C, 59.2; H, 6.13; N, 6.56. Found: C,62.31; H, 6.92; N, 6.

Condensation product of rifamycin B with di-n-propylmethylhydrazine: C HN O M.W. 868.01. Analysis: C, 63.65; H, 7.54;N, 4.84. Found: C, 64.14;H, 7.70; N,

Condensation product of rifamycin B with dirnethylethyl hydrazine: C H NO M.W. 825.95. Analysis: C, 62.53; H, 7.20; N, 5.08. Found: C, 61.92; H,6.34; N, 4.65

Condensation product of rifamycin B with triethyl hydrazine: C H N OM.W. 853.99. Analysis: C, 63.28; H, 7.43; N, 4.92. Found: C, 63.32; H,7.45; N, 4.62.

Condensation product of rifarnycin B with dipropylethyl hydrazine:C4'1'H6'1N3013, M.W. 882.03. Analysis: C, 64.00; H, 7.65; N, 4.76.Found: C, 63.70; H, 7.50; N, 4.58.

Condensation product of rifamycin B with diethyl-propyl hydrazine: C H NO M.W. 868.01. Analysis: C, N, 4.84. Found: C, 64.06; H, 7.86; N,

Condensation product of rifamycin B with diethylbutyl hydrazine:C47H67N3O13, M.W. 882.03. Analysis: C, 64.00; H, 7.65; N. 4.76. Found:C, 63.57; H, 7.85; N, 4.80.

Condensation product of rifamycin B with l-methylamino piperidine: C H NO M.W. 851.98. Analysis: C, 63.43; H, 7.21; N, 4.93. Found: C, 63.28; H,7.59; N, 4.64.

Condensation product of rifamycin B with l-ethylamino piperidine: C H NO M.W. 866.0. Analysis: C, 63.79; H, 7.33; N, 4.85. Found: C, 63.55; H,7.58; N, 4.80.

Condensation product of rifamycin B with l-propylamino piperidine: C4qH5N3O13, M.W. 880.02. Analysis: C, 64.14; H, 7.44; N, 4.77. Found: C,63.79; H, 7.70; N, 4.57.

Condensation product of rifamycin B with l-butylamino piperidine:C48H67N3O13, M.W. 894.02. Analysis: C, 64.48; H, 7.55; N, 4.70. Found:C, 64.15; H, 7.88; N, 4.38.

Condensation product of rifamycin B with 4-methylamino-morpholine: C H NO M.W. 853.97. Analysis: C, 61.85; H, 6.96; N, 4.92. Found: C, 61.31; H,7.25; N, 4.81.

Condensation product of rifamycin B with 4-ethylamino-morpholine: C H NO M.W. 867.99. Analysis: C, 62.26; H, 7.08; N, 4.84. Found: C, 61.50; H,7.39; N, 4.86.

Condensation product of rifarnycin B with 4-propylamino morpholine: C HN O M.W. 882.01. Analysis: C, 62.64; H, 7.19; N, 4.76. Found: C, 61.18;H, 7.67; N, 4.65.

Condensation product of rifamycin B with 4-buty1- amino morpholine: C HN O M.W. 896.03. Analysis: C, 63.00; H, 7.31; N, 4.68. Found: C, 61.92;H, 7.52; N, 4.40.

Condensation product between rifamycin B and tripropylhydrazine: C H N OM.W. 896.05. Analysis: C, 64.34; H, 7.76; N, 468. Found: C, 64.15; H,7.69; N, 4.53.

Condensation product between rifamycin B and diproyl-butylhydrazine: C HN O M.W. 910.07. Analysis: C, 64.66; H, 7.86; N, 4.61. Found: C, 64.23;H, 7.95; N, 4.57.

Condensation product between rifamycin B and dioutylmethyl-hydrazine: CH N O M.W. 896.08: Analysis: C, 64.34; H, 7.76; N, 4.68. Found: C,64.41; H, 7.90; N, 4.49.

Condensation product between rifamycin B and dibutylethyl-hydrazine: C HN3013, M.W. 910.07. Analysis: C,-64.66; H, 7.86; N, 4.61. Found: C,63.53; H, 7.65; N, 4.35.

Condensation product between rifamycin B and dibutylpropyl-hydrazine: CI-I N O M.W. 924.09. Analysis: C, 64.98; H, 7.96; N, 4.42.

We claim: I

1. A process for preparing rifamycin B hydrazides, which comprisesreacting, in a solvent and in the presence of a water-eliminating agent,rifamycin B with a hydrazine of the formula:

' HN-NB whereinlR is selected from the class consisting of hydroaccording to claim 1 in which the water eliminating agent isdicyclohexylcarbodi'imide.

.3. A process as in claim 2 whereinrifamycin B is reacted withfl-hydroxyethyl hydrazine.

N,4.54. Found: C, 64.33; H, 8.05;

4. A process as in claim 2 wherein rifamycin B is reacted withdipropylethyl hydrazine.

5. A process as in claim 2 Whereinrifamycin B is reacted with lmethylamino piper-idine.

6. A process as in claim 2 wherein rifamycin B is reacted withl-ethylamino piperidine.

7. A process as in claim 2 wherein rifamycin'B is reacted withl-propylamino piperidine. 8. A rifamycin B hydrazide selected from theclass consisting of the reaction products of rifamycin B and hydrazinesof the formula wherein R is selected from the class consisting ofhydrogen and lower alkyl, and NB is selected from the class consistingof amino, monO- and di-lower alkylamino, hydroxyethylarnino, anilino,lower alkyl-phenylamino, 4- pyridylcarbamyl, 4-pyridylamidino,l-piperidino, 4-morpholino, l-hydantoino and l-thioureido.

9. Rifamycin B fl-hydr-ox'y-ethyl-hydrazide. 10. Rifamycin Bdipropyl-ethyl-hydrazide. I 11. 1-(N-methyl-rifamycinamido)-piperidine.12. 1-(N-ethyl-rifarnycinamido) piperidine. l3. 1-(N-propyl-rifamycinamido) -piperidine. 14. 4- (N-ethyl-rifamycinamidomorpholine.

4-(=N-propyl-rifamycinamido)-morpholine. 16. 4- (N-butyl-rifamycinamido-morp'holine.

No references cited.

WALTER A. MODANCE, Primary Examiner.

1. A PROCESS FOR PREPARING RIFAMYCIN B HYDRAZIDES, WHICH COMPRISESREACTING, IN A SOLVENT AND IN THE PRESENCE OF A WATER-ELIMINATING AGENT,RIFAMYCIN B WITH A HYDRAZINE OF THE FORMULA:
 8. A RIFAMYCIN B HYDRAZIDESELECTED FROM THE CLASS CONSISTING OF THE REACTION PRODUCTS OF RIFAMYCINB AND HYDRAZINES OF THE FORMULA